The Nuclease Mechanism
There are two different mechanisms used by various nucleases to cleave the chemical bonds of the DNA or RNA polymer. The most common mechanism is one in which a water molecule is used to break the phosphodiester bond. This is called a hydrolysis reaction. Under most conditions the P-O (phosphorous and oxygen) bond of the DNA or RNA polymer is very stable, and the H2O molecule is not usually very reactive. However, nucleases that use the hydrolysis mechanism make the H2O reactive by removing one of the hydrogens to generate a highly reactive OH- (hydroxyl). The negatively charged OH- can then attack the P-O bond to cleave the polymer. An alternative mechanism used by some DNA repair endonucleases involves the initial cleavage of a C-O (carbon and oxygen) bond and subsequent P-O bond cleavage. This is called a lyase reaction and does not involve water.